Recent experiments in our laboratory have demonstrated that Coxsackie B 3 (CBS) virus induces IL-6 and IL-8 production in human cells. Infection of mice with CBS is associated with elevated levels of cytokines in heart tissue of wild type but not TLR4 or CD14 knockout mice. CBS, like other enteroviruses, has been shown to bind to the complement regulatory protein decay accelerating factor (DAF). In vitro data indicate, however, that expression of the Coxsackie and Adenovirus Receptor (CAR) is required for infection of cells with CBS. This cell surface protein is expressed in mouse embryos at approximately day 10, and expression decreases after birth in some organs, including the heart. However, pathologic examinations in humans and rodents indicate increased expression of CAR in subjects with myocardial disease. TLR4 and CD14 knockout mice develop lower levels of virus in their hearts suggesting a major role for TLRs in viral myocarditis. We plan to use our expertise in characterization of the study of TLRs and the CAR protein to define the interaction of TLR proteins, cytokines, and the CAR protein. We will test the hypothesis that the origin of myocarditis relates to: 1) infection of gut tissue (pancreatitis) by CBS, 2) upregulation of the CAR protein, and 3) increased virus in the heart. At the same time we will define the roles of CD14, and several TLRs (especially TLR2, TLR4 and TLR7) in the response to CBS and its regulation in the development of myocarditis. [unreadable] [unreadable]